Blood collection and separating devices have from time immemorial, spun down the whole blood in a container having its long axis oriented parallel, or mostly parallel, to the direction of the centrifugal force. Examples can be seen in, e.g., U.S. Pat. No. 4,012,325. There are several reasons for this orientation. One reason is that when centrifugal forces cease, there is a substantial distance of separation between the heavier red cells and the lighter serum, and at the same time, an interface between the two phases of reduced surface area. As a result, when serum is drawn off, there is less likelihood that the blood cells will redisperse into the lighter serum phase. To further prevent this undesired event, a gel of intermediate specific gravity is often used, to occupy the surface area between the two phases. The spinning of the container about the long axis insures that the depth of the gel that resists remixing after centrifuging, will be substantial.
Stated from an opposite point of view, it has not been considered feasible to spin such containers about one of the shorter axes. The reason is that the distance between the free surface of the separated serum, and its interface with the separated blood cells, becomes very short, with a concommittant large surface area at said interface. This in turn makes blood cell contamination of the serum as it is "poured off" or removed, more likely. Any attempt to use a gel to shore up such a large surface area interface is less likely to succeed, since the gel will have only a short depth to it to resist remixing. (The volume of the gel will be distributed primarily over that large surface area of the interface.)
However, the conventional approach has paid a price for these conclusions. The price is, that Phase separation takes a long time since it has to occur over the longest dimension of the liquid volume. For example, in a blood volume of about 2 mL, using a device similar to that described in the aforesaid '325 patent, the time of separation of the serum from the blood cells is on the order of 5.3 min when spinning at, e.g., 100 g's. It is true, of course, that such separation times are also a function of the centrifugal force applied--the greater the force (e.g., created by higher rpm values), the faster the separation. Thus, typically the forces that are used are well in excess of 1000 g's, as lower forces will cause unacceptable delay in the phase separation. But even at such higher forces, such as 1600 g's, the separation in a 2 mL volume container has not been generally possible in less than 30 sec. Most importantly, however, is a disadvantage that has now been discovered about such centrifugal forces: at the interface between the blood cells and the serum is a layer called the "buffy coat". Among other things, when formed at centrifugal forces in excess of 100 g's, the buffy coat has as an inseparable part thereof, leukocyte cells such as the lymphocyte cells, which contain useful DNA. If those cells could be drawn off, the DNA could be extracted. The problem has been that the phase separation that occurs using conventional containers and centrifuges therefor, insures that those lymphocyte cells are irretrievably mixed with the rest of the buffy coat. It will be readily apparent, therefore, that any attempt to speed up phase separation to less than one minute by drastically boosting the force of spinning, will completely interfere with the retrieval of the lymphocyte cells.
Therefore, prior to this invention there has been a substantial need for a blood phase separation method that allows faster phase separation and/or lower spinning forces, while at the same time somehow solving the high risk of remixing of the phases, noted above.
One approach to dealing with this need would be, of course, the provision of some mechanism that allows for ready withdrawal of the light serum phase from the container, before the centrifugal force is removed. This in turn will aid in retaining the unwanted blood cells in a capture zone of the container, during serum removal, since the centrifugal force will still be applied. In fact, a blood separator device has been proposed that allows serum removal from the container while spinning still occurs--it even occurs by increasing the spinning speed. The device in question is shown, for example, in Japanese Kokai 63/237368. A valve is provided closing off exit passageway from the container, it being spring biased so that it will open only when the centrifugal force is increased beyond the speed used during phase separation, e.g., from 3000 to 5000 rpm. Clearly, in such a device serum can be drawn off with a minimum of risk of red cells remixing with the serum being drawn. However, even in such a device, it was not considered that the "while--centrifuging" serum withdrawal would permit reorienting the device to spin about its short axis. Instead, the device once again insists on the conventional spin orientation wherein the phase separation must occur over the long axis of the container.
Another disadvantage of the device shown in the Japanese publication is that the valve will stay open as long as a high centrifugal force is applied, even in the absence of liquid flow. Clearly, a better construction is one in which the valve automatically closes after all serum is removed. The reasons are that a) failure to do so makes it possible that non serum components, if somehow loosened in the container, can also get out the valve, and b) the still open valve prevents other processing from being accomplished while spinning, on the blood cells remaining in the container. This disadvantage stems from the fact that the valve of this prior device operates only in response to centrifugal force, and NOT in response to the presence of liquid, e.g., serum, which is to be drawn off.
There has been a need, therefore, prior to this invention, for a two phase liquid separation method that will more promptly, and at slower speeds, achieve phase separation and automatic removal of the lighter phase, particularly when processing whole blood.